Ruthenium ammine complexes as electron acceptors for growth stimulation by plasma membrane electron transport

Ammineruthenium(III) complexes have been found to act as electron acceptors for the transplasmalemma electron transport system of animal cells. The active complexes hexaammineruthenium(III), pyridine pentaammineruthenium(III), and chloropentaammineruthenium(III) range in redox potential (E ₀) from 305 to –42 mV. These compounds also act as electron acceptors for the NADH dehydrogenase of isolated plasma membranes. Stimulation of HeLa cell growth, in the absence of calf serum, by these compounds provides evidence that growth stimulation by the transplasma membrane electron transport system is not entirely based on reduction and uptake of iron.     

Paleolimnological evidence for recent acidification of Big Moose Lake,Adirondack Mountains,N.Y. (USA)

Big Moose L. has become significantly more acidic since the 1950s, based on paleolimnological analyses of sediment cores. Reconstruction of past lakewater pH using diatom assemblage data indicates that from prior to 1800 to ca. 1950, lakewater pH was about 5.8. After the mid-1950s, the inferred pH decreased steadily and relatively quickly to about 4.6. Alkalinity reconstructions indicate a decrease of about 30 eq · l^-1 during the same period. There was a major shift in diatom assemblage composition, including a nearly total loss of euplanktonic taxa. Chrysophyte scale assemblages and chironomid (midge larvae remains also changed in a pattern indicating decreasing lakewater pH starting in the 1950s. Accumulation rates of total Ca, exchangeable and oxide Al, and other metals suggest recent lake-watershed acidification. Cores were dated using²¹⁰Pb, pollen, and charcoal. Indicators of watershed change (deposition rates of Ti, Si, Al) do not suggest any major erosional events resulting from fires or logging. Accumulation rates of materials associated with combustion of fossil fuels (polycyclic aromatic hydrocarbons, coal and oil soot particles, some trace metals, and sulfur) are low until the late 1800s-early 1900s and increase relatively rapidly until the 1920s–1930s. Peak rates occurred between the late 1940s and about 1970, when rates declined.The recent decrease in pH of Big Moose L. cannot be accounted for by natural acidification or processes associated with watershed disturbance. The magnitude, rate and timing of the recent pH and alkalinity decreases, and their relationship to indicators of coal and oil combustion, indicate that the most reasonable explanation for the recent acidification is increased atmospheric deposition of strong acids derived from combustion of fossil fuels.     

Influence of dose and route of administration of bovine follicular fluid and the suppressive effect of purified bovine inhibin (Mr 31,000) on plasma FSH concentrations in ovariectomized ewes

Ovariectomized Merino ewes were used to develop an in-vivo bioassay for purified bovine inhibin of Mr 31,000. Various doses (0.25, 0.5, 1 or 2 ml) of bovine follicular fluid, given either by the intravenous (i.v.) or intracarotid route (i.c.) resulted in significant linear dose-related suppression of plasma FSH and interval to maximum suppression. Control ewes (1.0 ml steer plasma) showed no significant change in FSH over the same period. Doses of 470 and 2590 U of pure inhibin given i.v. caused a significant suppression of FSH in plasma in all ewes. The in-vivo potency estimate of the high dose (2760 U, 1420-4690 fiducial limits) agreed well with the in-vitro assay of potency. There were no significant changes observed in mean plasma LH after treatment with the higher dose of pure inhibin. There were no rebound effects of treatment with bovine follicular fluid or pure inhibin on FSH concentrations above that of controls. It is concluded that the form of bovine inhibin of Mr 31,000, which is believed to be the predominant circulating form, is biologically active when administered in vivo.     

Comparison of the gonadal response of wild and laboratory field voles (Microtus agrestis) to different photoperiods

Weanling male and female field voles from laboratory stock and from the F1 generation of wild-caught animals were placed in a long (16L:8D) or short (6L:18D) photoperiod for 28 or 56 days. Both types of field vole showed the well-established effect of photoperiod upon sexual maturation, with animals in the long photoperiod having larger and more active gonads than animals in the short photoperiod. After 28 and 56 days laboratory stock females were more mature, sexually, and had a higher growth rate than did Wild F1 females. There was no difference between the two types of males at 28 days, but by 56 days laboratory stock males were more sexually mature and had a higher growth rate than did Wild F1 males. These differences between the two types occurred in the long and short photoperiods. There was no interaction between photoperiod and type of vole. The use of laboratory stock animals in experiments could lead to an incorrect assessment of the effect of photoperiod in the control of seasonal breeding in wild populations.     

Immuno-gold localization of α-L-arabinofuranosyl residues in pollen tubes of Nicotiana alata Link et otto

Immuno-gold labelling using a monoclonal antibody (PCBC3) with a primary specificity for -L-arabinofuranosyl residues was used to locate these residues in pollen tubes of Nicotiana alata grown in vivo. The antibody bound to the outer fibrillar layer of the pollen-tube wall: the inner, non-fibrillar wall layer was not labelled. Cytoplasmic vesicles (0.2 m diameter) were also labelled. The antibody may bind to an arabinan in the pollen-tube wall.     

A single amino acid substitution deregulates a bacterial lactate dehydrogenase and stabilizes its tetrameric structure

We have engineered a variant of the lactate dehydrogenase enzyme from Bacillus stearothermophilus in which arginine-173 at the proposed regulatory site has been replaced by glutamine. Like the wild-type enzyme, this mutant undergoes a reversible, protein-concentration-dependent subunit assembly, from dimer to tetramer. However, the mutant tetramer is much more stable (by a factor of 400) than the wild type and is destabilized rather than stabilized by binding the allosteric regulator, fructose 1,6-biphosphate (Fru-1,6-P2). The mutation has not significantly changed the catalytic properties of the dimer (Kd NADH, Km pyruvate, Ki oxamate and kcat), but has weakened the binding of Fru-1,6-P2 to both the dimeric and tetrameric forms of the enzyme and has almost abolished any stimulatory effect. We conclude that the Arg-173 residue in the wild-type enzyme is directly involved in the binding of Fru-1,6-P2, is important for allosteric communication with the active site, and, in part, regulates the state of quaternary structure through a charge-repulsion mechanism.     

Induction of siderophore activity in Anabaena spp. and its moderation of copper toxicity.

Growth of Anabaena sp. strain 7120 (in the absence of chelators or added iron) was inhibited by the addition of 2.1 to 6.5 microM copper and was abolished by copper concentration of 10 microM or higher. When the copper was chelated to schizokinen (the siderophore produced by this organism in response to iron starvation), the toxic effects were eliminated. Analysis of culture filtrates showed that the cupric schizokinen remains in the medium, thereby lowering the amount of copper taken up by the cells. Although this organism actively transports ferric schizokinen, it apparently does not recognize the cupric complex. Thus, Anabaena sp. is protected from copper toxicity under conditions in which siderophore is being produced. For cells grown in low iron, the accumulation of extracellular schizokinen was observed to parallel cell growth and continue well into stationary phase. The actual iron status of the organism was monitored by using iron uptake velocity as an assay. Cultures grown on 0.1 microM added iron were found to be severely iron limited upon reaching stationary phase, thus explaining the continued production of schizokinen. These data show that the siderophore system in Anabaena spp. has developed primarily as a response to iron starvation and that additional functions such as alleviation of copper toxicity or allelopathic inhibition of other algal species are merely secondary benefits.     

Rational construction of a 2-hydroxyacid dehydrogenase with new substrate specificity

Using site-directed mutagenesis on the lactate dehydrogenase gene from Bacillus stearothermophilus, three amino acid substitutions have been made at sites in the enzyme which we suggest in part determine specificity toward different hydroxyacids (R-CHOH-COOH). To change the preferred substrates from the pyruvate/lactate pair (R = -CH3) to the oxaloacetate/malate pair (R = -CH2-COO-), the volume of the active site was increased (thr 246----gly), an acid was neutralized (asp-197----asn) and a base was introduced (gln-102 - greater than arg). The wild type enzyme has a catalytic specificity for pyruvate over oxaloacetate of 1000 whereas the triple mutant has a specificity for oxaloacetate over pyruvate of 500. Despite the severity and extent of these active site alterations, the malate dehydrogenase so produced retains a reasonably fast catalytic rate constant (20 s-1 for oxaloacetate reduction) and is still allosterically controlled by fructose-1,6-bisphosphate.